RESUMO
Excitatory amino acid transporters (EAATs) represent a protein family that is an emerging drug target with great therapeutic potential for managing central nervous system disorders characterized by dysregulation of glutamatergic neurotransmission. As such, it is of significant interest to discover selective modulators of EAAT2 function. Here, we applied computational methods to identify specific EAAT2 inhibitors. Utilizing a homology model of human EAAT2, we identified a binding pocket at the interface of the transport and trimerization domain. We next conducted a high-throughput virtual screen against this site and identified a selective class of EAAT2 inhibitors that were tested in glutamate uptake and whole-cell electrophysiology assays. These compounds represent potentially useful pharmacological tools suitable for further exploration of the therapeutic potential of EAAT2 and may provide molecular insights into mechanisms of allosteric modulation for glutamate transporters.
Assuntos
Sistema X-AG de Transporte de Aminoácidos/antagonistas & inibidores , Sítios de Ligação/efeitos dos fármacos , Doenças do Sistema Nervoso Central/tratamento farmacológico , Transportador 2 de Aminoácido Excitatório/antagonistas & inibidores , Sistema X-AG de Transporte de Aminoácidos/química , Sistema X-AG de Transporte de Aminoácidos/genética , Animais , Sítios de Ligação/genética , Transporte Biológico/efeitos dos fármacos , Doenças do Sistema Nervoso Central/genética , Doenças do Sistema Nervoso Central/patologia , Biologia Computacional , Transportador 2 de Aminoácido Excitatório/química , Transportador 2 de Aminoácido Excitatório/genética , Humanos , Ligação Proteica/efeitos dos fármacos , Transmissão Sináptica/efeitos dos fármacos , Interface Usuário-ComputadorRESUMO
At the onset of a drug discovery program, the goal is to identify novel compounds with appropriate chemical features that can be taken forward as lead series. Here, we describe three prospective case studies, Bruton Tyrosine Kinase (BTK), RAR-Related Orphan Receptor γ t (RORγt), and Human Leukocyte Antigen DR isotype (HLA-DR) to illustrate the positive impact of high throughput virtual screening (HTVS) on the successful identification of novel chemical series. Each case represents a project with a varying degree of difficulty due to the amount of structural and ligand information available internally or in the public domain to utilize in the virtual screens. We show that HTVS can be effectively employed to identify a diverse set of potent hits for each protein system even when the gold standard, high resolution structural data or ligand binding data for benchmarking, is not available.
Assuntos
Avaliação Pré-Clínica de Medicamentos/métodos , Ensaios de Triagem em Larga Escala/métodos , Tirosina Quinase da Agamaglobulinemia/antagonistas & inibidores , Tirosina Quinase da Agamaglobulinemia/química , Indústria Farmacêutica , Antígenos HLA-DR/química , Antígenos HLA-DR/metabolismo , Humanos , Modelos Moleculares , Receptores Nucleares Órfãos/química , Receptores Nucleares Órfãos/metabolismo , Conformação Proteica , Inibidores de Proteínas Quinases/farmacologia , Fatores de Tempo , Interface Usuário-ComputadorRESUMO
We have developed a workflow to extract, separate, and semi-quantify bioactive oxysterols from mouse colon tissues and fecal matters using solid- and liquid-phase extractions, enzymatic and chemical modifications, and stable-isotope dilution LC/MS/MS. The method was applied to a dextran sodium sulphate (DSS)-induced mouse colitis model, which revealed that one particular dihydroxycholesterol (diOHC), 7α,25-diOHC, was significantly elevated in both colon tissue and fecal matters of mice with colitis compared to that in naïve mice. The extent of 7α,25-diOHC elevation was positively correlated with colitis severity.
Assuntos
Colite/induzido quimicamente , Colo/química , Modelos Animais de Doenças , Oxisteróis/isolamento & purificação , Animais , Cromatografia Líquida , Colo/patologia , Sulfato de Dextrana , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Oxisteróis/química , Espectrometria de Massas em TandemRESUMO
Here, we report a high-throughput virtual screening (HTVS) study using phosphoinositide 3-kinase (both PI3Kγ and PI3Kδ). Our initial HTVS results of the Janssen corporate database identified small focused libraries with hit rates at 50% inhibition showing a 50-fold increase over those from a HTS (high-throughput screen). Further, applying constraints based on "chemically intuitive" hydrogen bonds and/or positional requirements resulted in a substantial improvement in the hit rates (versus no constraints) and reduced docking time. While we find that docking scoring functions are not capable of providing a reliable relative ranking of a set of compounds, a prioritization of groups of compounds (e.g., low, medium, and high) does emerge, which allows for the chemistry efforts to be quickly focused on the most viable candidates. Thus, this illustrates that it is not always necessary to have a high correlation between a computational score and the experimental data to impact the drug discovery process.
Assuntos
Inibidores Enzimáticos/farmacologia , Isoenzimas/antagonistas & inibidores , Inibidores de Fosfoinositídeo-3 Quinase , Desenho de Fármacos , Ensaios de Triagem em Larga Escala , Simulação de Acoplamento Molecular , Estudos ProspectivosRESUMO
The synthesis and structure-activity relationships of a series of novel phenoxyphenyl diamine derivatives with affinity for both the histamine H(3) receptor and the serotonin transporter is described.
Assuntos
Diaminas/química , Antagonistas dos Receptores Histamínicos/química , Antagonistas dos Receptores Histamínicos/farmacologia , Receptores Histamínicos H3/efeitos dos fármacos , Inibidores Seletivos de Recaptação de Serotonina/química , Inibidores Seletivos de Recaptação de Serotonina/farmacologia , Antagonistas dos Receptores Histamínicos/síntese química , Humanos , Estrutura Molecular , Receptores Histamínicos H3/química , Inibidores Seletivos de Recaptação de Serotonina/síntese química , Relação Estrutura-AtividadeRESUMO
A series of novel 4-aryl-1,2,3,4-tetrahydroisoquinoline-based histamine H(3) ligands that also have serotonin reuptake transporter inhibitor activity is described. The synthesis, in vitro biological data, and select pharmacokinetic data for these novel compounds are discussed.